Project Summary Mitochondrial dysfunction in skeletal muscle has devastating consequences including muscle wasting, exercise intolerance, and insulin resistance. We have discovered that a novel, highly conserved protein is critical for maintenance of mitochondrial function and cellular energy homeostasis in yeast and mammalian cells. We have designated this protein Lifespan-associated Mitochondrial Stress-responsive 1 (Lms1). Our data from yeast support a model whereby Lms1 recruits components of the ubiquitin proteasome system to mitochondria to extract damaged proteins and present them to the proteasome for degradation. The purpose of this research is to determine the function and mechanism of Lms1 action in skeletal muscle using cultured muscle cells and Lms1 knockout mice. For Specific Aims 1 and 2, which span the K99 and R00 phases, the candidate will investigate the role of Lms1 in cultured muscle cells. Studies in Aim 1 will determine whether mammalian Lms1 recruits the ubiquitin proteasome system to mitochondria as part of a mitochondrial protein quality control system. Studies in Aim 2 will determine the nature of mitochondrial defects observed with Lms1 depletion in muscle cells. Completion of the sub- aims proposed during the K99 phase will provide the candidate with training in aspects of cellular and molecular biology necessary to independently complete the R00 phase. For Specific Aims 3 and 4, the candidate will determine the role of Lms1 at the mammalian organismal level. For Aim 3 (K99 phase), the candidate will examine an Lms1 knockout mouse for mitochondrial dysfunction in heart muscle and begin studies in skeletal muscle. For Aim 4 (R00 phase), the candidate will examine an Lms1 skeletal muscle- specific knockout mouse for mitochondrial dysfunction and consequences including exercise intolerance, muscle wasting, and insulin resistance. Experiments proposed in Aim 3 will provide the candidate with the training in mitochondrial physiology necessary to independently complete Aim 4 during the R00 phase. Collectively, these experiments seek to establish a mechanistic basis for Lms1 action in cultured muscle cells and to extend these findings to mice where they will be tested for physiologic relevance. These studies will provide novel insight into the regulation of mitochondria in skeletal muscle.